Prevalence and associated risk factors of malaria among febrile under-five children visiting health facilities in Ziquala district, Northeast Ethiopia: A multicenter cross-sectional study

Background Malaria is among the leading causes of mortality and morbidity among under five children in developing countries. Ethiopia has set targets for controlling and eliminating malaria through at-risk group interventions. However, the disease remains a serious public health concern in endemic areas like in Wollo, Northeast Ethiopia. Therefore, this study aimed to determine malaria prevalence, risk factors and parasite density among under five children in Ziquala district. Method A facility—based cross-sectional study was conducted in Ziquala hospital, and Tsitsika, Mishra and Hamusit health centers in Ziquala district, Northeast Ethiopia, from January 2022 to April 2022. The study enrolled a total of 633 under five children using a systematic sampling technique. A capillary blood sample was collected from each child to prepared thin and thick blood smears. Smears were then stained with 10% Giemsa and examined under light microscope. A pretested structured questionnaire was used to collect on socio-demographic data, parental/caregiver knowledge, and malaria determining factors. Bivariable and multivariable logistic regression analysis was done to identify factors associated with malaria. Result The overall prevalence of malaria among children visiting Ziquala district health institutions was 24.6% (156/633). Plasmodium falciparum, P. vivax, and mixed infection (both species) accounted for 57.1%, 38.5%, and 4.5% of the cases, respectively. Regarding to parasite load, moderate parasitemia was the most common, followed by low and high parasitemia with the proportion of 53.8%, 31.4% and 14.7% parasite density, respectively. Malaria infection was linked to irregular utilization of insecticide-treated bed nets (AOR = 5.042; 95% CI: 2.321–10.949), staying outside at night (AOR = 2.109; 95% CI: 1.066–4.173), and parents not receiving malaria health education in the past six months (AOR = 4.858; 95% CI: 2.371–9.956). Conclusion Malaria was prevalent among children under the age of five enrolled in the study. The local government should focus on regular insecticide treated net utilization, reducing the risk of mosquito bites while sleeping outdoors at night and increasing public understanding of malaria prevention and control through health education would also help to minimize the burden of malaria.


Introduction
Malaria is one of the most serious public health concerns and a life threatening parasitic disease caused by the Plasmodium species [1]. Malaria is the most common cause of death in children under the age of five years and pregnant women in developing countries [2][3][4]. Children are one of the most vulnerable groups because they are immunologically naive to malaria parasites. Malaria may cause as many as 10% of all deaths in children globally. Severe anemia, respiratory distress, and cerebral malaria are common complications in children with severe malaria [5].
Between 2000 and 2019, global malaria prevention and control activities were ramped up, and the World Health Organization African Region achieving significant success in reducing its malaria burden, particularly in Sub-Saharan Africa [6]. However, in tropical and subtropical areas of the world, malaria remains a major concern. The World Health Organization (WHO) 2021 malaria report indicates that the WHO African Region continues to bear the largest burden of malaria. In 2020, the African Region was accounted for 95% of all malaria cases (228 million); 96% of all malaria deaths (602 000); and 80% of all malaria deaths in the region were among children under the age of five. Malaria services were disrupted during the Covid-19 pandemic starting in 2020, adding to the region's malaria burden [7].
Despite strong investments to decrease malaria incidence rates over the past years, infection rates remain significant in sub-Saharan Africa. Children under the age of five in sub-Saharan Africa, where the great majority of malaria deaths occur each year, are particularly vulnerable to the disease [7]. Every day, over 1200 children die from malaria in this region, accounting for the majority of the continent's 631,000 child deaths [8]. With about 75% of Ethiopia's landmass estimated to be malarious, 68% of the country's population is at risk of malaria infection. According to Ethiopian Malaria Indicator Survey 2015, malaria is one of the top ten primary causes of morbidity. Plasmodium falciparum causes 60% of malaria cases, while the rest is caused by P. vivax [9].
To prevent and reduce malaria infections, several intervention activities such as the distribution of insecticide-treated nets (ITNs), indoor residual spraying (IRS), artemisinin-based combination therapy (ACT), and health information dissemination have been carried out in Ethiopia. These efforts notwithstanding, malaria remains a serious public health concern in endemic areas, particular among children under the age of five [10]. Ethiopia is implementing a malaria elimination program with the goal of eliminating the disease by 2030 [11]. To assess the progress of the program, the prevalence and determinants of malaria among vulnerable groups should be evaluated over time and in different areas.
Malaria is one of the top causes of morbidity and mortality in both adults and children under the age of five in Ziquala district in northeast Ethiopia, according to a district health office report. However, a study on the epidemiology of malaria in children under the age of five had not been conducted.Knowing the prevalence, parasite density, and determinants of malaria in children under the age of five in health facilities is critical for scaling up and designing effective intervention programs. Therefore, this study fills in an important knowledge gap by identifying potential risk factors for malaria infection among under five-year children in Ziquala District.

Study design, area, and period
A multi-site facility-based cross-sectional study was conducted in Ziquala hospital, and Tsitsika, Mishra and Hamusit health centers in Ziquala district, Northeast Ethiopia, from January 2022 to April 2022. Ziquala district is one of the eight districts of Wag Hemra Zone, which is located at a distance of 780 km from capital city of the country, Addis Ababa. The district has an altitude of 1462 meters above sea level. Its annual average rainfall and temperature are 255 mm and 22˚C, respectively [12]. The area's major and minor transmission seasons take place between September and December and between April and May, respectively [13]. Malaria infection is primarily due to P. falciparum. Subsistence farming and livestock breading are the main occupations in the area. Water conducive to mosquito breeding is available throughout the year because of rivers such as the Tekezie and Tela rivers. In the district, there is one primary hospital, five health centers, and fifteen health posts. All health facilities provide diagnostic and treatment services to the community.

Sample size determination and sampling technique
The sample size was calculated using a single population proportion formula, the 95% confidence limits (Zα/2 = 1:96), and 5% margin of error (d) with a maximum proportion of 50%.
Sample size = Z(α/2) 2 P (1-P)/ d 2 , (1.96) 2 � 0.221 (1-0.221)/ (0.05) 2 = 384. Accounting for a 10% non-response rate and implementing design effect of two, the final sample size was 633. Study health facilities were chosen by stratifying the district's health facilities as malaria-endemic and non-endemic based on their altitude. Then, among the resulting six health facilities in the district, four were selected for data collection by simple random sampling technique. The number of study participants to be enrolled at each health facility was determined based on the number of febrile under five children at each health facility during the same period (January 2021 to April 2021) of the previous year. Hence in this period, a total of 1240 febrile under five children were screened for malaria from Ziquala Hospital (482), and Tsitsika (376), Mishra (251) and Hamusit (131) health centers. Accordingly 246, 192, 128 and 67 febrile patients were included from Ziquala Hospital and Tsitsika, Mishra and Hamusit health centers, respectively. Finally, a systematic random sampling was applied to select study participants at each health facility.

Study population and eligibility criteria
The source population consisted of all children under the age of five who visited the designated health facilities during the study period. The study only included children whose parents or guardians signed a written consent form. Children who had undergone antimalarial chemotherapy 42 days before to data collection were excluded because the parasite is thought to be cleared from the blood within 42 days of treatment, and re-infection can happen after that.

Data collection
Socio-demographic and associated risk factors assessment. To collect data on sociodemographic and related risk factors for malaria, a standardized questionnaire was developed in English. It was then translated into Agewgna, the indigenous language. Data of socio-demographic and environmental were collected from their parents/caregivers using structured and pretested questionnaire by trained data collectors.

Laboratory data collection
Blood sample collection, processing, and examination. At Ziquala hospital, and Tsitsika, Mishra and Hamusit health centers laboratories; capillary blood was drawn aseptically from the children's fingers using a blood lancet. Thin and thick blood films were prepared and air dried. After that, the thin blood film was fixed with absolute methanol and allowed to air dry. Both thin and thick blood films were stained with 10% Giemsa stain for 10 minutes. The slides were then air dried after being washed with distilled water. Finally, laboratory technologists at the health facilities examined the stained slides. Thick smears were used to detect Plasmodium infections, while thin smears were employed for parasite species identification using an oil immersion objective.
To measure parasitemia, the asexual forms of the parasites, such as trophozoites and schizonts, were manually counted until 200 WBCs from thick smear using a tally counter. The number of parasites per microliter of blood was calculated using the formula below, with the patient's total leukocyte count assumed to be 8000/μl of blood. Parasite load is equal to the parasites counted per 200WBC X 8000WBC/μl blood. The parasitemia was then classified as low (<1000 parasites per microliter of blood), intermediate (1000-4999 parasites per microliter of blood) and high (>5000 parasites per microliter of blood) [14].

Data quality assurance
Data collectors were trained for two days by the investigators. The Giemsa stain was tested on known positive and negative malaria slides. During the microscopic examination, a slide was regarded as negative after 200 fields had been examined without finding of Plasmodium parasite by two laboratory technologists at each site. A colour atlas was used during microscopic examination. To assure quality of the microscopic examinations, all positive and 10% of the negative slides were re-examined by a third reader to remove discrepant result.

Data processing and analysis
Data were entered and analysed using SPSS version 22 after data collection. The prevalence rate and determinant factors were estimated using descriptive statistics. Bivariable and multivariable logistic regression models were used to find correlations with malaria infection. The multivariable logistic regression analysis included variables with a p-value of less than or equal to 0.25 in the bivariable regression. A p-value <0.05 was considered to determine statistical significance. Finally, the strength of the association between factors was determined using adjusted odds ratios (AOR) with a 95% confidence interval (CI).

Socio-demographic characteristics of study participants
A total of 633 febrile children under the age of five years old took part in the study, with a 100% response rate. Of the total participants more than half (53.9%) of individuals were males and 46.1% of them were females. Regarding to their age group [15], the highest number (33.2%) of children participated were in the age between 12 and 23 months. The majority of the children were come from rural areas (76.0%), while the rest from urban areas ( Table 1).

Prevalence of malaria and its density
The overall prevalence of malaria among under-five children in Ziquala district was 24.6% (95% CI = 21.1-28.1). Of these, 60.3% were males and 39.7% were females. Malaria prevalence in sex was not statistically significant difference (P = 0.065). Likewise, the higher number of malaria cases was detected a mong the age group of 12-23 and less than 12 months with the prevalence of 45.5% and 24.4%, respectively. Plasmodium parasite prevalence was higher in younger children, with a statistically significant difference (P = 0.001). The malaria parasite was shown to be more prevalent in children living in rural areas as compared to children in urban areas. Home village location was not associated with malaria infection (p = 0.455). In this study, the prevalence of P. falciparum and P. vivax was 57.1% and 38.5%, respectively, while that of mixed infection was 4.5% (Table 2).
Regarding to parasite density, moderate parasitemia was the most common, followed by low and high parasitemia, which accounted for 53.8%, 31.4%, and 14.7% individuals respectively. Males were found to have a higher prevalence of moderate malaria parasitemia than females, accounting for 57.1% of the total, however there was no statistical difference in parasite density (p = 0.683). Likewise, moderate parasitemia had the highest parasite density in the 12-23 age group, and there was statistical significance in parasitemia across age groups (p = 0.001). Moreover, there was a statistically differences (p = 0.001) between rural and urban residents, with 70.2% of rural residents having moderate parasitemia ( Table 3).
The highest numbers of children who participated were from Ziquala hospital (38.9%), while the lowest were from Hamusit health center (10.6%). There was also a considerable variation in the prevalence rate between the health facilities, ranging from 19.4% at Hamusit health center to 28.0% at Ziquala hospital ( Table 4).

Prevalence of malaria and knowledge of parents/caregivers
In this study, 72.0% of the parents/caregivers had no ability to read or write, whereas 28.0% had attained various levels of formal education. Of the total parents/caregivers, 91.9% believed

Factors associated with malaria infection
In this study, 94.0% of the respondents possessed an ITNs, and 96.1% had received IRS service in the previous 12 months. In terms of ITNs utilization, 58.0% of under-five children slept in an ITNs on a regular basis during the previous two weeks. In bi-variable logistic regression, the following variables were chosen and entered into the forward stepwise multivariable logistic regression model: stagnant water near the house, under five children sleeping under ITNs regularly for the last two weeks, number of under five children per household, staying outside at night, and health education on malaria in the previous six months.
In the final adjusted model, children under the age of five who did not sleep under an ITNs regularly for the last two weeks were more likely to be infected with malaria (adjusted OR  [AOR] = 5.042; 95% CI: 2.321-10.949) than those who did. Children who stayed outside at night had a 2.1-fold increased chance of contracting malaria than children who did not (AOR = 2.109, 95% CI: 1.066-4.173). In addition, children whose mother/caregiver had never received malaria health education in the previous 6 months also had a greater risk of infection (AOR = 4.858, 95%; CI = 2.371-9.956) than those whose mother/caregiver had received malaria health education (Table 6).

Discussion
The present study showed that malaria is a public health concern among children under five years old in the study area. The overall prevalence of malaria was 24.6% (95% CI = 21.1-28.1) among children visiting health institutions in Ziquala district. This result was comparable to a study conducted in Arsi Negele, Ethiopia (22.8%) [16], Arba Minch Zuria District, South Ethiopia (22.1%) [17] and Nigeria (22.6%) [18]. In contrast, the prevalence determined in this study showed a lower prevalence of malaria than a study conducted in Mali (35.4%) [19], Ghana (43.0%) [20] and Cameron (28.8%) [21]. The low prevalence of malaria parasites in this study could be due to seasonal variations, as the current study was conducted in a minor malaria transmission season. This difference might also be due to the difference in geographical variation, and malaria control and prevention programs implemented in the study areas. However, the prevalence of malaria in this study was higher than the prevalence determined by studies conducted in Wogera district, Ethiopia (8.7%) [15], Sherkole refugee camp, Ethiopia (3.9%) [22] and the national magnitude of malaria among under-five children (0.6%) [13], Sudan (18.0%) [23] and Uganda (19.5%) [24]. This could be due to the community's and responsible bodies' attention being diverted to the Covid-19 pandemic, which has resulted in less attention being paid to malaria prevention and control activities. Such an assertion is in accordance with the WHO global malaria report 2021 [7], which states that delays to malaria services during the Covid-19 pandemic had an additional influence on the malaria burden. In East Africa, the growth of antimalarial drug resistance is also a major problem [25,26]. This high prevalence of malaria in the study area proves that the efforts that have been made by the district, as well as the regional health office and other concerned bodies to control malaria in the study area, were not sufficient.
Plasmodium falciparum and P. vivax were the two species identified in the blood of the children, accounting for 57.1% and 38.5% of infections, respectively. This result is in line with the national Plasmodium species distribution [27] and other previous studies done in Ataye [28], Kombolcha [29] and Dembecha [30], Ethiopia. The higher dominance of P. falciparum over P. vivax can be explained by several factors. These include P. falciparum's high proliferation in the host cell, the parasite's capacity to infect all ages of red blood cells, the parasite's resistance to first line treatment [31] and the study area's has lowland climatic conditions, where P. falciparum is a widespread species in the lowlands.
In this study similar to a study done in Wogera district, Ethiopia [15], the number of children who had febrile illness and Plasmodium infection decreases with increasing age of a child. This is might be due to these children live in stable malaria transmission endemic areas and can develop age-related protective immunity as the result of continuous exposure to infective mosquito bites [32]. The majority of infected children had a moderate parasite density, followed by low and high parasite density, which accounted for 71%, 16% and 12.9% of malaria positive children respectively. This is consistent with an earlier study done in South Gondar, Ethiopia [33]. However, a high proportion of high parasite density was found in East Central Tanzania [34], while a high proportion of low parasite density was found in Sanja Town [35]. The immunological conditions, age category, and dietary status of the study participants could all have an impact on parasite density. By decreasing parasite load, acquired or adaptive immunity protects against clinical disease, morbidity associated with parasite density, and new infection [36].
Regarding to the health institutions, Ziquala hospital has the highest rate of malaria infection, followed by Tsitsika and Mishra health centers. Plasmodium falciparum was the predominant species found in all health facilities except Hamusit health center. This variability in the prevalence of malaria among different locations is in line with studies undertaken in Wolkite, Ethiopia [37], lowlands of southern Ethiopia [38] and Cameron [39]. This could be due to differences in malaria prevention and control interventions from one area to another. These variations could also be due to altitude differences and climate diversity which affect the reproduction of the Anopheles vector.
About 78.0% of the parents/caregivers had knowledge that mosquitoes are a vector for the transmission of malaria. This result was relatively lower than other studies done in Damot Gale, Wolayta Zone (85.0%) and Gondar (94.5%) Ethiopia [40,41]. However, knowledge of causes of malaria in this site was better than a study done in Gilgel Gibe, Ethiopia (70.1%) [42]. This difference might be due to endemicity of malaria and related efforts to control it among vulnerable population in the area. Among the study participants, 70.3% knew that stagnant water is a suitable breeding site for mosquitoes. Several studies in Ethiopia [15,22,33] have showed that malaria vector density and living close to a water body like a river or streams could be an important factors influencing malaria transmission.
When asked about malaria prevention measures, 73.0% of responders identified ITNs, whereas only 2.5% chose IRS. The knowledge of using ITNs (73.0%) and IRS (2.5%) for malaria prevention was lower than a study done in Damot Gale, Ethiopia [40]. According to WHO malaria control and elimination strategies, access to all interventions enhance reduction in malaria, including enhanced case management, scale-up of ITNs, IRS, early diagnosis and treatment, and environmental management [43]. The majority of the children (84.0%) sought treatment at least within 24 hours after the onset fever. Early detection and treatment, within 24 hours of the onset of symptoms are an essential element of malaria control. This finding was higher than the same figure in Damot Gale (71.7%) and Mandura (38.7%) Districts, Ethiopia [40,44]. This increased health-seeking behaviour within 24 hours could be due to the presence of good awareness towards prompt access to antimalarial drugs.
This study revealed that those children who did not reguraly use ITNs were 5.04 times more likely than those who did to be infected with the Plasmodium parasite It was similar with prior studies conducted in South Gondar and Arbaminch Zuria District, Ethiopia [17,33]. Insecticide-treated nets are an effective vector control method for preventing malaria transmission. When they use ITNs consistently, the risk of getting a mosquito bite might be decreased [45]. Therefore, regular ITNs usage must be enforced by local authorities in malaria endemic areas.
Children who stayed outdoors at night were more likely to be exposed from malaria infection compared to those that did not. Staying outside during the night showed a statistically significant association with malaria. This finding was in line with the previous studies done in Armachiho, in Dembia district and Sherkole refugee camp in Benishangul-Gumuz, Ethiopia [22,46,47]. This could be explained by exophagic-exophilic mosquito biting behavior [48]. Children whose mother/caregiver had never received malaria health education in the previous 6 months had a greater risk of infection than those whose mother/caregiver had received. This finding was also supported by the previous studies done in Uganda and Ghana [20,49].

Conclusion and recommendations
The prevalence of malaria in under-five children attending at selected health facilities of Ziquala district was high. The highest prevalence of malaria was found in those aged between 12 and 23 months old, and a higher proportion of moderate parasitemia was also observed in this study. Irregular use of bed net, staying outside at night and parents' not receiving malaria health education were the main correlates of malaria. The local government and other concerned bodies should give focus on regular ITNs utilization, infection associated with staying outdoors at night, environmental management and changing attitudes towards malaria prevention and control through health education to minimize the burden of malaria.